A Mycobacterial Systems Resource for the Research Community

Abstract

Functional characterization of bacterial proteins lags far behind the identification of new protein families. This is especially true for bacterial species that are more difficult to grow and genetically manipulate than model systems such as Escherichia coli and Bacillus subtilis. To facilitate functional characterization of mycobacterial proteins, we have established a Mycobacterial Systems Resource (MSR) using the model organism Mycobacterium smegmatis. This resource focuses specifically on 1,153 highly conserved core genes that are common to many mycobacterial species, including Mycobacterium tuberculosis, in order to provide the most relevant information and resources for the mycobacterial research community. The MSR includes both biological and bioinformatic resources. The biological resource includes (i) an expression plasmid library of 1,116 genes fused to a fluorescent protein for determining protein localization; (ii) a library of 569 precise deletions of nonessential genes; and (iii) a set of 843 CRISPR-interference (CRISPRi) plasmids specifically targeted to silence expression of essential core genes and genes for which a precise deletion was not obtained. The bioinformatic resource includes information about individual genes and a detailed assessment of protein localization. We anticipate that integration of these initial functional analyses and the availability of the biological resource will facilitate studies of these core proteins in many Mycobacterium species, including the less experimentally tractable pathogens M. abscessus, M. avium, M. kansasii, M. leprae, M. marinum, M. tuberculosis, and M. ulcerans. IMPORTANCE Diseases caused by mycobacterial species result in millions of deaths per year globally, and present a substantial health and economic burden, especially in immunocompromised patients. Difficulties inherent in working with mycobacterial pathogens have hampered the development and application of high-throughput genetics that can inform genome annotations and subsequent functional assays. To facilitate mycobacterial research, we have created a biological and bioinformatic resource (https://msrdb.org/) using Mycobacterium smegmatis as a model organism. The resource focuses specifically on 1,153 proteins that are highly conserved across the mycobacterial genus and, therefore, likely perform conserved mycobacterial core functions. Thus, functional insights from the MSR will apply to all mycobacterial species. We believe that the availability of this mycobacterial systems resource will accelerate research throughout the mycobacterial research community.

Spatiotemporal localization of proteins in mycobacteria

“…In our recent effort to understand the subcellular organization of mycobacteria ( Judd et al, 2021 ), we systematically tagged over 1,000 highly conserved M. smegmatis ( Msm ) proteins with the fluorescent protein Dendra ( Gurskaya et al, 2006 ) and generated a comprehensive microscopy dataset as part of the Mycobacterial Systems Resource (MSR) ( Judd et al, 2021 , example images and the primary analysis of the dataset available on https://msrdb.org ). While these images are valuable, they have limitations that prevent us from fully exploiting the information they contain.…”

Structure-Aware Mycobacterium tuberculosis Functional Annotation Uncloaks Resistance, Metabolic, and Virulence Genes

“…(N) Mycobacterial core genes. Genes identified recently as part of the mycobacterial core genome ( 67 ) and annotated as hypothetical proteins (according to Table S3 of the recent work) are listed, along with their updated annotation and its source(s). Download .…”

“…Our updated annotation provides function for 34.4% (45/131) of genes with hypothetical function identified in a recent systems resource as broadly conserved across mycobacteria ( 67 ) ( Data Set S3 contains the full set). Mycobacterial core genes annotated include functions well established experimentally, such as essential component of the mycobacterial transcription initiation complex RbpA ( https://gitlab.com/LPCDRP/Mtb-H37Rv-annotation/-/blob/master/features/Rv2050.tbl ) and others not evident from extant literature but of potential clinical relevance, like the host-directed effector function inferred for Rv3909 ( https://gitlab.com/LPCDRP/Mtb-H37Rv-annotation/-/blob/master/features/Rv3909.tbl ).…”

Myrtus Communis Leaf Extract - A Source of Secondary Metabolites Exhibiting Anticancer and Antimycobacterial Activities

“…7B) that compared to the control, the bio lm formation of M. smegmatis was drastically reduced in the wells treated with ½ MIC of extract than the wells wherein the cells were treated with ¼ MIC of M. communis extract. Unlike M. smegmatis, which forms the bio lm at the liquid-air interface [23], S. aureus forms the bio lm at the solid surface, which was di cult to be photographed.…”